Colony Morphology Lab Report

This lab experiment was conducted in order to identify and confirm the presence of three types of organism within an unknown broth culture. The organism that were being tested fall under the categories of gram-positive, gram-negative paracolons, and gram-negative coliform. Gram positive organism are known to have thick cell wall made up of peptidoglycan, which are cross-link sugar chains with peptide bonds (Carson 13). Gram-positive bacteria are found within the phylum Firmicuites (Slonczewski & Foster 94). While gram-negative, have thin layers of one or two peptidoglycan cell wall, this type of bacteria are found within the phylum Proteobacteria (Slonczewaki & Foster). The gram positive and negative characteristic derived from a staining technique that was developed by Hans

Christian Gram (Carson 13). This technique help applies a differential stain to an organism in order to help classify or determine the organism of interest base on cell wall composition. Diverse type of antibiotics interact differently with different groups of organism, this is pertinent information when it comes to prescribing antibiotics. Organism of a particular gram stain are known to share certain characteristic. This is of great importance in clinical setting, because this technique can help reduce ambiguity during the diagnosis process. In essence, to know the type of gram bacteria you are looking for within a sample can help narrow the choices down to a particular type of organism. Just like the differential gram stain, there are different culturing techniques that can help in the isolation of a particular organism. For all intended purposes, the streak plate technique is used to help isolate high organism colony concentrations to individual colony (Carson 19).

Individual colonies are better to work with because the organism can be aseptically transferred to another media, to create a pure culture. This helps narrow down specifically the type of organism of interest (Carson 19). During the culturing process, there are a lot of information that can help deduce the type of organism that are cultured. Some of the characteristic that are used in the identification of cultured organism are known as colony morphology (Carson 20). Colony morphology take in consideration the following characteristic of the culture: Size (small, medium, large), pigment (coloration), optical properties (opaque, translucent, transparent), surface (shiny, smooth, and mucoid) form (circular, irregular, and rhizoid), margin (lobate, undulate, filamentous or serrate), and elevation (flat, raised, umbonate, or convex) (Carson 20). In a more microscopic level, cellular characteristic may apply to the individual cell within a colony, otherwise known as cellular morphology. The agar plate were used during this experiment have selective and differential properties that help to differentiate between organism. A selective media will actively select for or against a particular

organism. A selective media select for an organism base on the availability of nutrients or a chemical that favor the growth of a type of organism while preventing the growth of another organism. Differential media allows the organism to interact with it in order to produce a distinguishable result. The combination of both media were used during this experiment to determine the type of organism that are available within the unknown. Another aspect that was taken into consideration was the pH level of the medium. During the metabolic process of the organism within the media, acid or base byproduct can result from this process (Carson 58). A pH indicator will help identify the effect of the organism to the media; this in return will help determine the type of organism activities that are occurring on the media. The specific medium that were used for this experiment were as followed: TSA, blood agar, MacConkey, Eosin Methylene Blue Levine (EMB), Salmonella-Shigella, Mannitol Salt Agar (MSA), Triple Sugar Iron (TSI), and Citrate. The TSA plate test was used to study the morphology of the cultured organism from the unknown broth mixture. The blood agar was use to determine the hemolytic properties of the gram-positive bacteria within the unknown mixture, this is the ability of an organism to metabolize red blood cells. Blood agar is considered a differential medium. The MacConkey test is known for its ability to select for gram-negative bacteria and prevention of the growth of gram –positive bacteria. This is useful in isolating the two type of gram-negative bacteria from the gram-negative within the unknown sample. The MacConkey agar is considered a selective and differential media. The EMB test help specify between the fermentation nature of gram-negative bacteria. Coliform is the category of organism that can break down lactose while paracolon is the category denote to organism that cannot metabolize lactose. The EMB media differentiate between those metabolic characteristic by allowing itself to be distinguish through the acid byproduct that are produce because of the metabolism of lactose. This organism is consider as a elective and differential media The SS plate was used to validate the presence Shiglla and Salmonella within the unknown culture, because this medium allows for the growth of those types of organism and prevent the growth of others. This medium is considered a selective and differential plate. The MSA plate is a selective and differential plate that select for Staphylococcus genus, because those bacteria are able to grown within the high salt concentration of MSA. Other organism are inhibited because they cannot grow on such high concentration of salt. Carbohydrates and pH indicator help indicate the metabolism of glucose within the media, low pH during high metabolic process. The triple sugar iron test is a differential medium that select bacteria base on their ability to ferment glucose, sucrose, and lactose, the gas produce during this process and the reduction of hydrogen sulfide (Carson 68). The citrate test will test the bacteria ability to produce the enzyme citrase that metabolize sugar. In the presence of citrase the mediun turns pH will increase and thus allows the chemical bromothymol blue to distinguish between bacteria base on that aspect. Within this experiment the hypothesis is develop base on expected outcomes of the different media and test. Given the prior knowledge of the type of bacteria that were available within the unknown (gram positive, gram negative, parcolon, coliform), the hypothesis was composed as followed: the gram-positive bacteria that are potentially present within the culture are Staphylococcus aureus, Staphylococcus epidermis, Bacillus subtillus, Streptococcus pyogenes, Streptococcus pneumonia. For the gram-negative paracolons, the hypothesis proposed for the testing of the presence of Salmonella enterica or Yersinia enterocolitica. For the gram-negative coliform, the hypothesis was proposed for the presence of Klebsiella pneumonia, Escherichia coli.